秋水仙素加倍玉米单倍体胚性愈伤组织的初步研究
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1广东省农业科学院作物研究所 / 广东省农作物遗传改良重点实验室,广州 510640;2 四川农业大学玉米研究所,成都 611130

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基金项目:

广东省重点领域研发计划项目,2018B020202008;广东省农业科学院“十四五”农业优势产业学科团队建设项目,202115TD;广东省级乡村振兴战略专项,粤财农【2020】100号


Preliminary Study on Colchicine Doubling Haploid Embryogenic Callus of Maize
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1Institute of Crops,Guangdong Academy of Agricultural Sciences/Guangdong Key Laboratory of Crop Genetic Improvement, Guangzhou 510640;2 Maize Research Institute of Sichuan Agricultural University,Chengdu 611130

Fund Project:

Guangdong Provincial Key Research and Development Program, 2018B020202008,

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    摘要:

    以玉米单倍体胚性愈伤为遗传转化受体,结合化学加倍方法,可以在较短时间内获得纯合稳定的个体,加快转基因植株的纯合时间。本研究基于二倍体幼胚胚性愈伤诱导方法,获得玉米单倍体愈伤,并分析了单倍体愈伤检出率,不同继代次数后的褐化率和死亡率,以及二倍体愈伤与单倍体愈伤的生长速率等重要指标。同时,采用0.005%-0.03%秋水仙素处理单倍体愈伤,分析处理后单倍体愈伤的死亡率,成株后的散粉率,大花药(花粉活力强)比率,结实率以及结实大于5粒的植株比率等指标。研究结果表明,基于改进的二倍体愈伤诱导方法,能够正常诱导出单倍体胚性愈伤,单倍体愈伤检出率为4.2%,与诱导系MT-1对自交系18-599(红)的诱导率相当。单倍体愈伤的褐化率和死亡率均明显高于同继代次数下的二倍体愈伤,且继代3次后的单倍体愈伤褐化率和死亡率最高。以愈伤直径为参考,单倍体愈伤的生长速率低于二倍体愈伤,在生长早期可作为判断单倍体和二倍体愈伤的重要参考指标。经不同浓度和时间的秋水仙素处理后,单倍体愈伤的散粉率、大花药比率、结实率以及结实大于5粒植株比率随处理浓度和时间的增加而提高,而存活率和小花药(活力较低或无活力)比率随处理浓度和时间的增加而下降。由此表明,秋水仙素处理可提高玉米愈伤成株后的散粉率和结实率,结实率的提高又取决于大花药比率的提高。本文综合分析了玉米单倍体愈伤诱导及加倍过程中的诸多因素,筛选出0.01%秋水仙素处理12h(处理编号:B12-2,成活率58%,结实率29%)和0.02%秋水仙素处理3h(处理编号:C3-2, 成活率59%,结实率31%)作为玉米单倍体愈伤加倍的较优处理模式。

    Abstract:

    Using maize haploid embryogenic callus as genetic transformation receptor and combining with chemical doubling method, homozygous individuals whenever hosting a transgenic element might be obtained in a short time. This study attempted to obtain haploid callus from maize using diploid embryogenic callus induction method, followed by analyzing the detection rate of haploid callus, browning rate and mortality after different subculture times, as well as diploid callus and haploid callus growth rate and other important indicators. The haploid callus was treated with 0.005%-0.03% colchicine, and the mortality rate of haploid callus, powder dispersal rate after adult, ratio of large anther (strong pollen vigor), seed setting rate and plant ratio of more than 5 seeds were analyzed. The results showed that induction of haploid embryogenic callus was applicable using the improved method of diploid callus induction. The detection rate of haploid callus is 4.2%, similar to the induction rate of 18-599(red) by MT-1 lines. Both the browning rate and mortality of haploid callus were significantly higher than diploid callusof the same time of subculture, and both the browning rate and mortality rate of haploid callus reached the highest level after three inductions. With callus diameter as reference, the growth rate of haploid callus was lower than t diploid,. , which could be used as an key reference indicator to distinguishhaploid and diploid callus in the early growth period. After being treated with colchicine of different concentrationsfor different period of time, the pollinating rate, rate of large anther, seed setting rate and ratio of plants with more than 5 seeds of haploid callus increased as the concentration increases and the time prolongs, and survival rate and small anther (of poor or no viability), on the other hand, drops as the concentration increases and the time prolongs.. The results showed that colchicine treatment could enhance the pollinating rate and seed setting rate of maize callus in adult plants where higher seed setting rate was dependent on higher rate of large anther.. Collectively, this paper includes analysis of multiple factors in the process of callus induction and doubling of maize haploid. And screened out 0.01% colchicine treatment for 12h and 0.02% colchicine treatment for 3h) as better modesfor doubling of maize haploid callus.

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李高科,陈琦,孟鑫,等.秋水仙素加倍玉米单倍体胚性愈伤组织的初步研究[J].植物遗传资源学报,2021,22(6):1606-1614.

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  • 收稿日期:2021-04-08
  • 最后修改日期:2021-07-17
  • 录用日期:2021-07-19
  • 在线发布日期: 2021-11-04
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